Inhibition of the menin-MLL protein-protein interaction is a promising new therapeutic strategy for the treatment of acute leukemia carrying MLL fusion (MLL leukemia). We describe herein our structure-based design, synthesis and evaluation of a new class of small-molecule inhibitors of the menin-MLL interaction (hereafter called menin inhibitors). Our efforts have resulted in the discovery of highly potent menin inhibitors, as exemplified by compound 42 (M-89). M-89 binds to menin with a K d value of 1.4 nM and effectively engages cellular menin protein at low nanomolar concentrations. M-89 inhibits cell growth in the MV4;11 and MOLM-13 leukemia cell *
Targeting the menin-MLL protein−protein interaction is being pursued as a new therapeutic strategy for the treatment of acute leukemia carrying MLL-rearrangements (MLLr leukemia). Herein, we report M-1121, a covalent and orally active inhibitor of the menin-MLL interaction capable of achieving complete and persistent tumor regression. M-1121 establishes covalent interactions with Cysteine 329 located in the MLL binding pocket of menin and potently inhibits growth of acute leukemia cell lines carrying MLL translocations with no activity in cell lines with wild-type MLL. Consistent with the mechanism of action, M-1121 drives dose-dependent down-regulation of HOXA9 and MEIS1 gene expression in the MLL-rearranged MV4;11 leukemia cell line. M-1121 is orally bioavailable and shows potent antitumor activity in vivo with tumor regressions observed at tolerated doses in the MV4;11 subcutaneous and disseminated models of MLL-rearranged leukemia. Together, our findings support development of an orally active covalent menin inhibitor as a new therapy for MLLr leukemia.
Breast cancer is a leading cause of death in women and its metastatic spread to distant organs is responsible for over 90% of these deaths. Circulating tumor cells (CTCs) implicated in the metastatic process occur when cancer cells undergo cytoskeletal and phenotypic transitions that enhance motility, then penetrate into the blood circulation. While CTC dissemination has typically been considered as the spread of single cells, recent evidence suggests that collective migration results in polyclonal circulating clusters that are the main drivers of metastasis. To study circulating clusters we have developed a dual microfiltration system employing two stages of antigen-independent mechanical cell isolation. As whole blood is passed through the system, circulating clusters are trapped by the first stage while single CTCs are retained by the second stage. Characterization experiments using cell lines demonstrated a capture efficiency of 87.1% for clusters and 85.7% for single cells on their respective microfilters. This system was used to investigate circulating cells in the metastatic MMTV-PyMT syngeneic mouse breast cancer model. Both clusters and single CTCs were detected in mice with histological evidence of lung metastases (14-16 weeks old), but not in mice that had not yet developed metastatic tumors (<12 weeks old). Immunofluorescence assays were established to determine expression of CD24, CD90, cytokeratins, CD45 and DAPI. Tumor cells were identified as nucleated cells that expressed epithelial cytokeratins, but not the leukocyte antigen CD45. While PyMT cells with a CD90+/CD24+ stem cell phenotype comprise only ~1.5% of tumors, they were observed in > 50% of circulating clusters. The presence of cancer stem cells was correlated to cluster size, with smaller clusters containing a greater percentage of CD90+/CD24+ cells. Isolated clusters will be further analyzed to quantify expression levels of cancer stem cell markers in each individual constituent cell. The analysis of circulating clusters may provide crucial insight into the mechanisms of metastatic disease and provide the means to discover novel targets for therapeutic intervention.
Citation Format: Ramdane Harouaka, Gabrielle Hodges, Allison David, Rishika Polasani, Spencer Freeman, Katherine Sanchez, Kaitlin Harvey, Max S. Wicha. Circulating clusters in breast cancer express cancer stem cell phenotypes [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 4776. doi:10.1158/1538-7445.AM2017-4776
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